The multifunctional nature of mast cells is clearly demonstrated through their involvement in both innate and adaptive immune responses. Recent insight into the various functions of mast cells has revealed that these innate immune effectors possess the dual ability to kill microbes and to modify classical adaptive immune responses. Members of the cathelicidin family of antimicrobial peptides (AMPs) are expressed in mast cells and at sites of epithelial injury. The localization of cathelicidin peptides, the trigger for their extracellular release, and their capacity to modify inflammatory responses all suggest that cathelicidins play an important role in the capacity of mast cells to fight infection. We have demonstrated that mouse mast cells use cathelicidin to fight infections caused by skin pathogens. Unfortunately, very little is known on human mast cell antimicrobial function and no studies describing cathelicidin AMP activity in human mast cells are available. The major goal of this proposal is to investigate how human mast cells regulate cathelicidin expression to fight infections caused by skin pathogens. This investigation proposes to: Specific Aim 1: Identify and define human mast cell cathelicidin. a. Identify signals that regulate expression of human mast cell cathelicidin. b. Define cathelicidin processing in human mast cells. Specific Aim 2: Investigate the antimicrobial activity of human mast cell cathelicidin. a. Investigate the role of Toll-like receptor activation in human mast cell antimicrobial activity. b. Evaluate the antimicrobial activity of processed human mast cell cathelicidin peptides. c. Ascertain the contribution of cathelicidin to human mast cell antimicrobial activity. Specific Aim 3: Establish an animal model to test the antimicrobial activity of human mast cell cathelicidin. a. Generate extracorporeal cultures of mast cells that express human or mouse cathelicidin. b. Reconstitute mast cell-deficient mice with cultured mast cells. c. Challenge mice intradermally with bacteria. This exploratory project will bring together investigators with unique and complementary areas of expertise in order to enhance our understanding of the role of cathelicidin AMPs in skin immunity. This knowledge has the potential to lead to the establishment of new therapeutic targets and strategies to combat infections by human pathogens that use the skin as a portal of entry. We believe that this proposal is particularly responsive to the goals of PA-06-181 and the exploratory developmental grant mechanism. PUBLIC HEALTH RELEVANCE Mast cells defend themselves and keep pathogens under control through their ability to produce antimicrobial peptides such as beta-defensins and cathelicidins. We have shown in preliminary experiments that mouse mast cells sense bacteria through receptors, which alert specific enzymes that prepare cathelicidin antimicrobial peptides to kill invading pathogens. This proposal seeks to unravel our understanding of this innate immune pathway in human mast cells with the ultimate goal of enhancing antimicrobial peptide activity.